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Catherine Burkhart1,2, Geraldine Paszkiewicz3, Loretta Gawron2, Rachael Kohrn2, Andrei Purmal1,4, Elizabeth Repasky3, Carl Morrison3, Angela Omilian3, Andrei Gudkov1,3,4 and Katerina Gurova3
1Cleveland BioLabs, Inc (Buffalo, NY, USA), 2Buffalo BioLabs, LLC (Buffalo, NY, USA), 3Roswell Park Cancer Institute (Buffalo, NY, USA), 4Incuron, LLC (Buffalo, NY, USA)
Background: CBL0137 represents a novel class of small molecules, Curaxins, that
simultaneously activate p53 and inhibit cancer-associated stress response
pathways, such as NF- B and HSF-11. CBL0137 antitumor activity has been
established in a broad range of tumor models both by oral and intravenous
administration1. The effects of CBL0137, culminating in tumor cell death, are
mediated by the sequestering of the FACT (FAcilitates Chromatin Transcription)
complex on chromatin thereby inhibiting its activity1. FACT is a transcription and
replication factor (composed of two subunits, Structure Specific Recognition
Protein (SSRP1) and suppressor of Ty 16 (SPT16)), which is involved in
transcription of genes with highly ordered chromatin structure as well as in
replication and mitosis1-3. FACT is expressed during early embryogenesis and in
undifferentiated progenitors and stem cells of adult tissues while protein levels
of both FACT subunits are almost undetectable in differentiated cells and
tissues2. FACT is expressed in several tumor types compared to equivalent
normal tissues. In addition, FACT positive tumors have an aggressive malignant
phenotype as illustrated by a correlation of FACT expression with high grade,
the development of metastatic disease and worse overall survival4. Thus, FACT
represents a potentially important target for cancer therapy. In particular, a high
proportion of pancreatic cancers express FACT as demonstrated by ~60% of
tested pancreatic tumors staining positive for the SSRP1 subunit (n=56)4.
Furthermore, 30% of these cases expressed a very high level of SSRP1.
1Gasparian et al (2011) Science Translational Medicine 3:95ra74 2Garcia et al (2011) Oncotarget 2:783-96 3Reinberg and Sims (2006) Journal Biological Chemistry 281:23297-23301 4Garcia et al (2013) Cell Reports (in press) http://dx.doi.org/10.1016/j.celrep.2013.06.013
METHODS: • Patient-derived pancreatic ductal adenocarcinoma (PDA) xenografts
were obtained under an approved IRB protocol and continually propagated in
SCID mice for use in efficacy experiments.
• Immunohistochemistry: TMAs containing 56 distinct PDA samples were
stained for SSRP1 and samples were scored for both % of positive cells and
intensity by trained a histopathologist.
• Pilot CBL0137 efficacy studies: SCID mice (n=5/treatment group; 2
tumors/mouse) were implanted with 2-5 mm3 of the designated patient PDA. ~10
days following implantation (tumor volumes <200 mm3) treatment commenced.
For oral administration, mice received vehicle or 30 mg/kg CBL0137 by gavage
following a 5 days on/2 days off schedule for 4 weeks. For iv studies, mice
received either vehicle or 90 mg/kg CBL0137 via the tail vein once per week up to
4 weeks. Tumors were measured 2-3 times per week using digital calipers. Tumor
volume was calculated using standard equation mm3 = L x W2/2 where L=
longest dimension and W is measured perpendicular to L. Mice were followed
until total tumor burden of mouse reached ~ 2000 mm3, all controls completed the
study or 29 days, whichever came first. Data is presented a mean fold tumor
growth by normalizing the tumor volume on Day X by that of Day 1. Comparisons
between treatment groups were performed by ANOVA.
• Combination efficacy studies: These studies were performed as described
above with the following modifications. Treatment commenced when the mean
tumor volume per group was ~25-50 mm3. Mice were treated with 80-90 mg/kg
CBL0137 iv (1/wk) in the presence or absence of 40 mg/kg gemcitabine
administered ip every 4th day (Q4d) for 4 weeks. Mice were followed until at least
one tumor per mouse reached 1000 mm3 or up to 90 days from start of treatment.
Normal CBL0137
NF B
p53
RNA - pol II
RNA
NF B RNA - pol II
p53 p53
p53 p53
p53 p53
p53 p53
p53 p53
p53 p53
RNA - pol II
nucleosome
CBL0137
Figure 1. Mechanism of action of CBL01371
Normal
PDA#13590 PDA#13756 PDA#15010
PDA#10978
Figure 3. SSRP1 Expression levels in patient derived PDA Xenografts
Figure 4. Effect of 30 mg/kg CBL0137 administered po (5 days on/2 days off for 4 weeks) on patient-derived PDA xenograft. Data is presented relative to Day 1 tumor volumes. * P<0.05, ANOVA.
PDA#12914
* * * * * *
* * * *
Oral Administration of CBL0137
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CBL0137
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CBL0137
IV Administration of CBL0137
Figure 5. Effect of 90 mg/kg CBL0137 administered iv (once per week for up to 4 weeks) on 4 independent patient-derived PDA xenografts that were propagated through mice upon collection from patients. Studies for individual tumors ended when all control tumors reached the designated tumor volume endpoint. Data is presented relative to Day 1 tumor volumes. * P<0.05, ANOVA. N=10 tumors per treatment group.
Conclusions: • FACT represents a potential target for cancer therapy due to its level of expression compared to normal tissue,
association with an aggressive malignant phenotype, metastatic disease and poor overall survival (Garcia et al
(2013) Cell Reports (in press) http://dx.doi.org/10.1016/j.celrep.2013.06.013). It is found in a large proportion of
PDA.
• CBL0137, which targets FACT, caused a 49-76% reduction in FACT positive PDA xenografts derived from
patient tumors when administered orally daily or intravenously once per week.
• CBL0137 produced a similar antitumor effect to gemcitabine and Abraxane against a gemcitabine/Abraxane-
sensitive PDAs during the treatment period and appeared to enhance the antitumor effect of Abraxane and
delay the recurrence of tumors in combination with gemcitabine at a dose, in which it was ineffective itself.
Acknowledgments: This work was funded in part by an STTR grant awarded by the NCI (Buffalo BioLabs, LLC,
KG and CB) and a grant from Incuron, LLC (KG).
Sub-optimal CBL0137 dose significantly delays recurrence of gemcitabine-regressed patient-derived PDA xenografts
0 10 20 30 40 50 60 70 80 90 0
10 20 30 40 50 60 70 80 90
100 vehicle gemcitabine only CBL0137 only GEM + CBL0137
days from start of treatment
Perc
en
t su
rviv
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PDA#13756
P=0.0153
Figure 7. Effect of 80 mg/kg CBL0137 on survival time of PDA-bearing mice in the presence or absence of gemcitabine.
Vehicle80 mg/kg CBL0137,
iv, 1/wk40 mg/kg
gemcitabine, ip, Q4dCBL0137 +
gemcitabine
Median survival time (days)
16 17 54 78
IV Administration of CBL0137 +/- Gemcitabine or +/- Abraxane
Figure 6. Effect of 80-90 mg/kg CBL0137 administered iv (1/wk for 4 weeks) ± 40 mg/kg gemcitabine (GEM) ip (Q4d for 4 weeks) or ± 30 mg/kg Abraxane (Abx) iv (D1-5) on patient derived PDA tumor growth.
-1 1 3 5 7 9
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PDA#13590
vehicle GEM CBL0137 (90) GEM/CBL0137
Last CBL0137 dose
Last GEM dose
0 2 4 6 8
10 12 14 16 18 20
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Days from start of treatment
PDA#13590
vehicle Abx CBL0137 (90) CBL0137/Abx
Last Abx dose
Last CBL0137 dose
PDA# 13756
0 5
10 15 20 25 30 35 40 45 50 55 60 65
0 5 10 15 20 25 30 35 40
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Days from start of treatment
vehicle CBL0137 (80) Abx CBL0137/Abx
Last Abx dose
Last CBL0137 dose
SSRP1 Expression levels in PDA Tissue Microarray Samples
SSRP1 Negative (<1)
SSRP1 + (>1)
High SSRP1
(>4)
Figure 2. TMAs including 56 PDA samples were stained for SSRP1 and scored based on % positive cells and intensity4.
BBL
Buffalo BioLabs, LLC
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